Icahn School Of Medicine At Mount Sinai

NIH Research Projects · FY 2025 · 2025-07

Project Summary Small animal models are ideal to study diverse infectious and inflammatory diseases with mice being a widely used model system due to the well-characterized immune response and availability of reagents. Yet, the drawbacks of the mouse animal model are that it does not mimic many diseases due to the lack of clinical symptoms, differences in immune activation, and non-susceptible to infection of many pathogens. In fact, ferrets, Guinea pigs, and hamsters are a more suited animal model for numerous infectious pathogens including bat coronaviruses (SARS-CoV and SARS-CoV-2), influenza virus, and respiratory syncytial virus, respectively; yet, the lack of reagents that define the immune and inflammatory responses in ferrets, Guinea pigs, and hamsters prevent their utilization to comprehensively understand disease pathology and effectiveness of therapeutics and vaccines. Thus, there is a severe reagent gap for monoclonal antibodies against immune and inflammatory markers of ferrets, Guinea pigs, and hamsters to use these animal models in infectious disease studies. The main objective of the R24 application is to create a Repository of Monoclonal Antibodies (RoMA) for use in small animal models for evaluating infectious and inflammatory disorders fulfilling a major reagent gap in the scientific community. We hypothesize that reagents that monitor the immune response in hamster and Guinea pig model systems will be widely used in infectious disease model systems to predict disease progression and effectiveness of vaccines and therapeutics in human disease. A panel of monoclonal antibodies will be generated upon completion of the following aims: Aim 1: Develop antibodies targeting hamster and guinea pig surface markers on the respective immune cells. The hamster has become an excellent model for respiratory pathogens including SARS-corona viruses, Rift Valley fever, and Clostridium difficile; while the Guinea pig animal model provides insight into numerous infection and transmission models such as influenza virus and herpesviruses. Aim 2: Identify and generate monoclonal antibodies to hamster and guinea pig immune cell activation markers. We plan to identify and generate monoclonal antibodies to activated immune cell immune markers of hamster and Guinea pigs. These activated biomarkers will consist of previously characterized human and mouse orthologs that likely function in immune regulation. We expect to generate a total of 50 monoclonal antibodies (25 anti-hamster and 25 anti-Guinea pig proteins) over the 5-year grant period that will be available to the scientific community. These antibodies will be essential for evaluating the immune and inflammatory responses in the hamster and Guinea pig animal models.

NIH Research Projects · FY 2025 · 2025-07

PROJECT SUMMARY The experiences and lessons learned from the COVID-19 pandemic geared our interests towards investigating the mechanism of action of different vaccine platforms and vaccination strategies that aim to induce long-lasting protective systemic and mucosal immune responses to prevent disease and transmission via the mucosal route. Dr. Weina Sun co-developed a Newcastle disease virus vector (NDV)-based COVID-19 vaccine that can be used either as an injectable inactivated whole virion vaccine or a live intranasal vaccine, the latter of which can elicit mucosal immune responses as a “stand-alone” vaccine platform. In naïve mice, a 2-dose regimen of intranasal NDV-based vaccine induced a high level of mucosal IgA antibodies. In mice that were pre- vaccinated with mRNA-LNP COVID-19 vaccines via the intramuscular route, a third intranasal NDV booster also substantially enhanced nasal wash antigen-specific IgA. By closely examining samples obtained from different anatomical mucosal sites in mRNA-LNP vaccinated mice without the NDV booster, spike-specific IgA were detected in nasal wash, intestinal lavage as well as vaginal lavage, among which Intestinal lavage contained the highest levels of IgA. Dr. Jennifer Gommerman is an immunologist with special interests studying mucosal B lymphocytes. In humans, she developed methods to detect salivary antibodies. While SARS-CoV-2 infection induces high levels of spike-specific salivary IgG and IgA, salivary IgG are also abundantly found after intramuscular 2-dose mRNA vaccinations. Intriguingly, some of the mRNA-vacccinated individuals developed and maintained modest salivary secretory-IgA that were resistence to decay after a 2-dose mRNA vaccine. Given that both in mice and humans, mucosal antigen-specific IgA were detected after mRNA-LNP vaccination, we hypothesize that mRNA-LNP vaccines can prime de novo mucosal B cells that are close to the administration sites, which are able to recirculate to other mucosal surfaces. Consequently, a further mucosal booster vaccine, such as the intranasal live NDVvaccine, would be able to recall pre-exisiting memory B cell responses. We will follow two specific aims to test our hypothesis, utilizing mRNA-LNP vaccine and NDV vaccine encoding the spike of Beta variant as a proof of concept: (Aim 1) To examine temporal and spatial dynamics of antigen-specific IgA and IgG secreting B cells in the mucosa-associated lymphoid tissues after mRNA and NDV vaccination in mice. (Aim 2) To track circulation of antigen-specific mucosal B cells in photoconvertible Kikume mice after mRNA prime followed by NDV vaccination or infection by flowcytometry. The proposed work entails innovative approaches and mouse models to test our hypothesis. We believe the outcomes of the study can inform the mechanism of action of not only mRNA-based and NDV-based SARS-CoV-2 vaccines, but also the same types of vaccines against other respiratory or enteric viral pathogens, in which mucosal antibody responses plays a critical role to mitigate virus infection and transmission.

NIH Research Projects · FY 2025 · 2025-07

PROJECT SUMMARY Opioid overdose-related deaths have claimed over 100,000 lives in 2022, necessitating a focus on biomarkers of recovery and relapse risk. As addiction progresses, the motivational control over drug use is posited to shift from being goal-directed (i.e., dependent on the drug's hedonic value and the ventral striatum and prefrontal cortex) to habitual (i.e., cue-triggered, via the dorsal striatum). Rooted in preclinical studies, the evidence for this cortico-striatal basis in human drug addiction is scarce. The available computational evidence suggests goal- directed control impairments that are associated with shorter abstinence and relapse in alcohol and methamphetamine use disorders. No study to date has inspected the neurobiology of motivational control, nor its putative fluctuations during recovery, in opioid use disorder. This K99/R00 application intersects naturalistic, longitudinal, and computational methods to inspect the contribution of goal-driven behaviors to opioid (heroin and fentanyl) addiction severity and recovery, encompassing craving and relapse. In the K99 phase, Aim 1 will track motivational control daily (using individual smartphones) for eight weeks during inpatient heroin addiction treatment (with and without fentanyl use), offering the candidate the opportunity to train in naturalistic and computational methods. Here, we expect impairments to be evident in early treatment but to normalize as a function of abstinence. Aim 2 will use the expected dynamic shifts in motivational control to inform changes in craving and relapse, tracked daily by ecological momentary assessments. The candidate will learn to harness longitudinal and predictive models for translational insights. We expect goal-directed control decreases to predict elevations in craving and relapse events. Aim 3 in the R00 phase will use a novel functional magnetic resonance imaging task that estimates choice under drug and nondrug contexts to naturalistically inspect motivational control cortico-striatal engagement in an independent sample of inpatients with opioid use disorder (inclusive of heroin and fentanyl). The candidate will gain expertise in translational neuroimaging methods. We expect lower goal-directed control to be associated with lower prefrontal cortex activity, evident especially in a drug cue context, in the patient group compared to controls. This proposal will allow the candidate to gain proficiency in the naturalistic, longitudinal, and computational study of drug addiction. The candidate will be guided by an excellent mentorship team: Drs. Rita Z. Goldstein (drug addiction neurobiology and its naturalistic study), Nathaniel Daw (computational modeling of motivational control), Katie Witkiewitz (longitudinal assessments of addiction treatment outcomes), Dr. Nelly Alia-Klein (clinical assessment and treatment of drug addiction), Hung- Mo Lin (consultant; biostatistical support), and Vincenzo Fiore (consultant; model-based neuroimaging), and the state-of-the-art resources at the Icahn School of Medicine at Mount Sinai. The innovative proposal and the carefully crafted training plan will propel the candidate to an impactful and independent research career, ultimately yielding insights into improving treatment outcomes for people with drug addiction.

NIH Research Projects · FY 2025 · 2025-07

The mission of the predoctoral Integrated Training in Pharmacological Sciences Program (ITPSP) at the Icahn School of Medicine at Mount Sinai (ISMMS) is to develop highly skilled biomedical scientists who will advance drug discovery through rigorous and ethical research, leveraging interdisciplinary approaches in pharmacology. Although submitted as a new application, this program builds on a 25-year legacy of pioneering quantitative training in pharmacology, emphasizing its importance for all PhD candidates in the biomedical sciences. We intend to appoint trainees to the T32 for two years, typically in their 2nd and 3rd years of PhD training, as they transition from classroom learning to full-time laboratory research. We aim to continue advancing innovative training that prepares students to become tomorrow’s leaders in pharmacology and to address the urgent need for new approaches in drug development. A cornerstone of ITPSP is its newly designed curriculum, which encourages students to harness quantitative methods with basic principles of molecular pharmacology to tackle biological concepts central to disease mechanisms and drug discovery. In our revamped coursework, students apply quantitative approaches through active learning and problem-solving exercises. Foundational biological concepts are taught in a disease context such as cardiovascular disease and metabolic disorders, in order to enhance engagement and translational insight. Student training is further reinforced through activities that emphasize experimental rigor, transparent reporting, and data reproducibility, which are integrated throughout the curriculum, dissertation committee meetings, and trainee evaluations. ITPSP trainees also benefit from a range of complementary activities that enhance the research experience, including journal clubs, Works-in-Progress seminars, and an annual retreat focused on career development, as well as alumni events that provide valuable networking opportunities. Another innovation of our program is its commitment to real-world experiences through internships in the pharmaceutical industry and Mount Sinai’s tech transfer office, among many other related activities. These opportunities, typically conducted during trainees’ 3rd or 4th years, foster industry relationships and broaden professional networks. The program is led by three experienced PIs: Program Director Dr. Avner Schlessinger and Co-Directors Drs. Eric Sobie and Ming-Ming Zhou. Each brings complementary expertise—Dr. Schlessinger in computational chemistry and transporter pharmacology, Dr. Sobie in quantitative systems pharmacology and graduate education, and Dr. Zhou in academic and industry-based drug discovery. ITPSP provides a supportive training environment that implements data-driven practices and thoughtful mentorship strategies to promote trainee well-being, retention, and success. In summary, ITPSP stands at the forefront of innovation, offering an evidence-informed and rigorous research training environment that prepares the next generation of pharmacology leaders to redefine drug development paradigms.

NIH Research Projects · FY 2025 · 2025-07

SUMMARY The research project focuses on understanding and modulating the plasticity of Th17 cells, specifically their ability to transdifferentiate into Treg cells (TregexTh17), a process with potential implications for autoimmune diseases, organ transplants, and malignancies. New insights indicate that AMP-activated protein kinase (AMPK) activity is highly involved in T cell plasticity. Yet, uncovering the biochemical mechanism employed by AMPK to drive TregExTh17 generation remains a significant challenge in the field, primarily due to the lack of appropriate technical approaches. We have developed a set of fluorescent kinase sensors capable of measuring the activity signatures of various kinases. These sensors employ diverse chemical approaches, offering a broad spectrum of colors ranging from UV to NIR. These sensors (i) are based on the specific substrate peptide sequence and are thus highly selective for the kinase/substrate pair of interest, an approach well-established in biosensing, (ii) are non toxic and thus suitable for live cell use, (iii) display rapid reactivity, and (iv) minimally interfere with the native cellular environment, preserving T cell functions. In this application, we propose a combinatorial approach using these cutting-edge, highly sensitive, and selective fluorescent biosensors to decipher the AMPK activity signatures responsible for driving Th17-to-Treg plasticity (TregExTh17) (Aim 1), and (ii) harnessing this plasticity through developing Phosphorylation Targeting Chimeras (PHICS), molecules designed to bring AMPK in proximity to key substrates involved in this plasticity, and phosphorylate/activate them as necessary to promote TregExTh17cells (Aim 2). After the R21 project, we will have developed a comprehensive fluorescent toolbox for monitoring AMPK kinase activity signatures in T cells and their relationship with function. By establishing connections between these signatures and their impact on Th17-to-Treg plasticity, we aim to unveil the translational potential of this innovative approach in deciphering the mechanisms governing this plasticity. Furthermore, we anticipate that developing a novel class of small molecules capable of selectively modulating substrate phosphorylation will strongly impact all contexts in which modulating immune responses is desired. These achievements will generate crucial data for R01 applications. 1

NIH Research Projects · FY 2025 · 2025-07

Project Summary Prostate cancer rates are elevated among World Trade Center (WTC) responders, and some cases have unfortunately recurred and progressed to aggressive disease. While our team has conducted one of the first molecular profiling of WTC-related prostate tumors, current studies rely on bulk-tissue analyses, which overlook the contributions of specific cell types and spatial interactions in the tumor microenvironment (TME). This knowledge gap limits our ability to understand the mechanisms driving recurrence and impedes the development of targeted therapies. To address this challenge, the objective of this application is to uncover the molecular and cellular mechanisms driving prostate cancer recurrence in individuals exposed to World Trade Center (WTC) toxins. By leveraging spatial transcriptomics and multi-omics analyses, we will identify key regulatory pathways that differentiate recurrent tumors from those in remission in the spatial context of their TME. Our central hypothesis is that prostate cancer recurrence is driven by spatially localized dysregulation of genetic pathways and cellular interactions, e.g., DNA repair pathways and aberrant tumor-immune interactions specific to certain regions within the TME. The research team with complementary expertise in cancer epidemiology, genome technology, and cancer multi-omics will leverage several key innovations in this research project—including the use of (i) the Mount Sinai WTC tumor biobank, (ii) cutting-edge spatial transcriptomic technologies, and (iii) integrative computational pipeline for bulk/single-cell data—to resolve molecular drivers underlying the recurrence of WTC-related prostate tumors at spatial and cellular definitions. The technical feasibility and commitment of the research team are demonstrated by the strong preliminary data demonstrating our completion of Visium HD spatial profilings on one recurrent vs. non-recurrent WTC-related prostate tumor, which will be expanded to 20 primary WTC-related prostate tumors that recurred vs. achieved long-term remission in this study. The two Aims are: Aim 1. Identify Spatial TME Patterns associated with Recurrence in WTC-Related Prostate Tumors Aim 2. Determine the Genomic and Epigenomic Regulatory Drivers of Recurrence in Prostate Cancer This project is significant because it addresses a critical knowledge gap in understanding the recurrence of prostate cancer, particularly in WTC responders. By identifying spatially localized molecular mechanisms driving recurrence, the study has the potential to inform the development of targeted therapies, ultimately improving outcomes for WTC-exposed prostate cancer patients and broader patient populations at risk for aggressive, recurrent disease.

NIH Research Projects · FY 2025 · 2025-07

Project Summary/Abstract The opioid epidemic has continued to be a major public health issue within the United States, contributing to ~100,000 overdose deaths. Opioid abuse contributes to more than just the more often focused addiction related behaviors. Long-term heroin exposure has recently been linked to the acceleration of neurodegenerative disease-like changes. Utilizing bulk RNA sequencing, we measured the gene expression signature within the dorsal striatum in human heroin users and aged matched controls. In addition to replicating previous findings relevant to behaviors related to the effects of opioids, we also identified a large set of genes that were altered related to mitochondrial oxidative phosphorylation and the electron transport chain. These genes, which were predominantly downregulated, were also enriched in KEGG ontology pathways related to several neurodegenerative diseases such as Alzheimer’s, Parkinson’s, and Huntington’s. My project aims to investigate the casual role heroin self-administration has on the mitochondrial transcriptional profile in the rodent dorsal striatum following heroin exposure using RNA sequencing. Additionally, I will use in-vivo and in-vitro models to elucidate the impact of opioid exposure on mitochondrial morphology and function. To that end, I will leverage immunohistochemistry to visualize perturbations on the mitochondrial network and use Seahorse assay to measure changes in oxygen consumption rate of mitochondria in cells to assess ATP production efficiency. Lastly, leveraging immunohistochemistry and functional strategies used in my cell culture and rodent model, I will assess mitochondria characteristics in post-mortem dorsal striatum tissue of heroin users. Altogether, this project will allow me to gain significant training in bioinformatics, RNA sequencing, metabolic assays, immunohistochemistry, and behavioral assays that will provide a significant foundation for my future development as an independent researcher to answer questions that might start from studies of the human brain to subsequently interrogate underlying neurobiological mechanisms and behavior in translational models.

NIH Research Projects · FY 2026 · 2025-07

Despite an unprecedented growth in Crohn's disease (CD) therapies it has remained consistent that each new therapy is less effective for patients that failed a previous therapy leaving many patients refractory to treatment. While CD therapies seemingly encompass distinct therapeutic mechanisms, all target the inflammatory response supporting that medically refractory CD involves mechanisms not directly related to inflammation. The near universal success of autologous stem cell transplantation (SCT) in refractory CD provides a rare cohort to understand therapeutic mechanisms that exist outside of the traditional CD treatment paradigm. In our study of CD patients that received a SCT we strengthened previous associations between inflammatory macrophages and refractory disease but further suggested a refractory phenotype is acquired through dysfunction of hematopoietic stem cells (HSC) and progenitors that reinforce differentiation of aberrant myeloid cells unable to support intestinal healing and rendering immunotherapy ineffective. As such, the central goal of this proposal is to understand differences in hematopoietic progenitors associated with refractory CD and how hematopoietic progenitor dysfunction determines clinical outcomes. This proposal will advance our understanding of CD therapeutic mechanisms through (Aim 1) profiling of HSC populations associated with refractory CD and (Aim 2) characterizing HSC functions associated with refractory CD and therapeutic outcomes. In Aim 3 we will explore a poorly defined intestinal HSC niche important to our long-term goal of understanding how myeloid progenitors support intestinal populations that reinforce refractory disease or promote healing.

NIH Research Projects · FY 2025 · 2025-07

PROJECT SUMMARY The individuals who were involved in the rescue and recovery efforts (responders) of the World Trade Center (WTC) tragedy were exposed to a complex mixture of airborne pollutants, and the health impact has been profound. Health conditions found to be associated with the WTC exposure are continuing to evolve. Since we are now over two decades from the event, it is imperative to evaluate how WTC exposure may impact the aging process and age-related disorders. There is emerging evidence that 9/11 exposure has an effect on age-related health outcomes, including frailty. Frailty increases with age and is defined as a clinical syndrome characterized by a decrease in one’s reserve and ability to maintain homeostasis across organ systems, and it has been linked to poor health outcomes. A World Trade Center Clinical Frailty Index (WTC-CFI) was developed to study frailty in this unique population and has been associated with high WTC exposure. As this vulnerable population ages, it is therefore critical to prioritize interventions to reduce modifiable risk factors that are potentially on the path towards frailty. One such modifiable risk factor is obesity. Given more than 80% of responders are overweight or obese and an elevated BMI has been associated with frailty, this is an important area of study. Our proposal aims to expand the understanding of the role of obesity on the trajectory of frailty and to determine the differences in objective physiologic and metabolic measurements in frail vs. non-frail obese responders to guide future evidence-based lifestyle interventions. We will achieve this with the following aims: 1) Determine the effect of obesity on frailty over time in the WTC GRC using the previously developed WTC CFI while controlling for other known, age-related risk factors; 2) Elucidate, for the first time, the role of obesity on frailty in a subset of frail and non-frail obese general responders from the Mount Sinai WTC Clinical Center of Excellence Healthy Steps for Older Adults (HSOA) intervention by comprehensively assessing differences in body composition, resting metabolic rate, substrate selection, serum biomarkers, dietary factors, and fitness measures to assess strength, aerobic capacity, and balance. Our proposal builds upon strong prior research and will be a crucial step in developing a greater understanding of both frailty and obesity and their complex relationship within this cohort that has already experienced a significant environmental exposure. Without this work, it would be challenging to create evidence-based lifestyle interventions for responders aimed at intervening on obesity as a modifying factor on the path towards frailty.

NIH Research Projects · FY 2025 · 2025-07

Project Summary/Abstract: Targeted therapies for the treatment of monogenetic obesity are essential because typical lifestyle interventions and standard anti-obesity medications are largely ineffective as they do not correct the specific genetic defect causing abnormal energy balance. The leptin pathway is the key regulator of body weight through control of appetite and energy expenditure. In particular, the severe insatiable hunger experienced by patients with leptin pathway disorders leads not only to extreme obesity, but the unrelenting drive to seek food also causes substantial distress for patients and their caregivers. While therapies have been developed for treating genetic disorders affecting the proximal portion of the leptin pathway (LEP, LEPR, POMC, PCSK1, and BBS1-22), there are no treatments for loss-of-function LOF) variants of the melanocortin-4 receptor gene (MC4R), which cause melanocortin obesity syndrome (MCOS). In various population and cohort studies, 1-6% of patients with severe, early onset obesity are found to have MC4R LOF variants, making MCOS the most common cause of genetic obesity. Brain-derived neurotrophic factor (BDNF) is a downstream mediator of MC4R signaling and therefore may serve as a specific target for MCOS treatment. We propose repurposing a well-understood and commercially available attention-deficit hyperactive disorder (ADHD) medication, atomoxetine, for the treatment of MCOS, because of animal and human studies showing that this drug induces endogenous BDNF levels. We hypothesize that atomoxetine will increase hypothalamic BDNF levels, leading to weight loss through improved anorectic signaling downstream of the abnormally functioning MC4R. To test our hypothesis, we propose to conduct a phase 2 randomized, placebo controlled, crossover trial in 20 patients with MCOS age ≥ 6 years. The primary outcome measure will be change in BMI (expressed as the percentage of the 95th percentile BMI for age/sex). Additional measures will include percent body fat and visceral fat area by bioelectrical impedance analysis, resting energy expenditure by indirect calorimetry, dietary intake by food frequency questionnaire and 24 hour recall, hyperphagia score, hunger level, satiety level, hemoglobin A1c, lipid panel, liver function tests, blood pressure, heart rate, and ADHD symptoms. Serum and plasma BDNF and genetic variants in atomoxetine metabolism enzymes will be assessed and correlated with weight changes. This pilot clinical trial will provide valuable data on the safety and efficacy of atomoxetine for treating MCOS, and the data will be used to guide the design of a future phase 3, multicenter, randomized clinical trial.

NIH Research Projects · FY 2025 · 2025-07

Abstract The US has recently experienced an alarming increase in the prevalence of allergic diseases. Allergic disease is characterized by an Immunoglobulin E (IgE) mediated immune response to common environmental antigens, with pathological manifestations ranging from rhinitis to life-threatening anaphylaxis. Under normal physiological conditions, production of IgE is tightly regulated by mechanisms affecting IgE- producing lymphocytes. A critical aspect of this regulation involves signaling through the B cell antigen receptor (BCR), which consists of a variable antigen-binding, membrane-bound immunoglobulin (mIg) and an invariant Ig⍺/Igβ signaling module. Unlike the canonical IgM BCR, the IgE BCR carries out robust, antigen-independent signaling. Autonomous activity of the IgE BCR induces IgE B cells towards developmental outcomes that restrict affinity maturation and militate against the emergence of IgE lymphocytes in long-lived or memory compartments. The aberrant production of allergen-reactive IgE can induce fatal anaphylaxis. Despite the potentially severe consequences of IgE dysregulation for human health, available data on ligand-independent IgE BCR signaling comes exclusively from murine models. Limited data suggests a conserved role for autonomous IgE BCR signaling pathways in human IgE B cells. Differences in IgE biology between humans and mice, however, highlight a need for further investigation in human model systems. Importantly, humans express two isoforms of membrane-bound IgE (mIgE), of which one is entirely absent in mouse. Human mIgE isoforms are differentiated by the length of their respective extracellular membrane-proximal domain (EMPD) primary sequences. The EMPD plays a key role in BCR assembly, suggesting a function for divergent EMPDs in modulating BCR signaling properties. I hypothesize that human mIgE isoforms give rise to IgE BCRs with distinct ligand-independent signaling properties. To address this hypothesis, I will perform a series of cell- based and molecular assays to characterize the ligand-independent signaling properties of human IgE BCR isoforms (Aim 1). In parallel, I will use a single particle analysis cryo-EM pipeline to obtain high-resolution structures of both human IgE BCR isoforms (Aim 2). Insights arising from this research will contribute to an understanding of how the IgE BCR regulates IgE responses in human health and disease, with implications for the development of strategies for managing conditions associated with IgE dysregulation. In parallel, this work will afford an exceptional training opportunity, supporting the development of expertise in structural biology, mass spectroscopic proteomic analysis, and immunobiology.

NIH Research Projects · FY 2026 · 2025-07

PROJECT SUMMARY The misuse of opioid painkillers targeting the -opioid receptor (MOR) continues to result in record numbers of overdose deaths, underscoring the pressing need to identify safer analgesics, as well as effective therapies for opioid use disorder (OUD). Although efficacy with (e.g., preferable drugs that can sufficiently engage pain pathways to provide therapeutic relief while leading to lower side effects through reduced engagement of their mediating circuits. Understanding not free from liabilities, several MOR small-molecules that exhibit low- agonism for relevant G protein subtypes (e.g., buprenorphine and oliceridine) are powerful analgesics i mproved safety profiles compared to other opioids used clinically (e.g., morphine) or commonly abused fentanyl). This supports the emerging paradigm in the field that low-efficacy MOR agonists may be candidates for developing howlow-efficacy agonists activate MOR-mediated G protein pathways differently from high-efficacy opioid drugs is essential for informing the design of safer opioid therapeutics. Achieving this understanding requires an unprecedented level of spatial resolution of both the conformational dynamics and kinetics of ligand-specific MOR-Gi1 coupling and activation, which cannot currently be obtained by a single technique. Instead, it necessitates the integration of data from both experimental and computational sources. Our team is uniquely positioned to tackle this challenge using an innovative Bayesian inference framework to integrate comprehensive conformational ensembles from physics-based enhanced molecular dynamics simulations with state-of-the-art single-molecule imaging data collected at unprecedented time resolution both inpurified protein preparations and in living cellsby long-standing experimental collaborators. This integration will yield more accurate atomistic information regarding ligand-specific MOR-G protein conformations and their kinetic relationships, from which we can derive accurate predictions about the unique attributes of low-efficacy opioids. Experimental validation of these predictions will not only help us achieve the overarching goal of elucidating the mechanistic basis of opioid efficacy but also bolster the architectures and inform optimal training of guidelines for developing novel chemotypes in the pursuit of safer opioid artificial intelligence medications.

NIH Research Projects · FY 2025 · 2025-07

Immediately after the 9/11 terrorist attack, concerns were raised about the short and long term effects of exposure to the World Trade Center (WTC) dust cloud. Responders and residents were exposed to a complex mixture of toxic chemicals that included multiple known and suspected human carcinogens which could increase cancer risk. Over the years, an increased risk in overall cancer incidence, and specifically of prostate, thyroid, melanoma and tonsil cancers has been observed. To understand the biological reasons for the observed cancer increase, in 2015 we established at MSSM the cancer tissue biobank (CTB), with CDC/NIOHS support. Currently 472 tumor tissue samples from responders have been collected, characterized and stored. Research on the cancer samples have significantly moved forward the field; for prostate, our own study on cancer tissues suggests the possibility of a biologically more aggressive cancer due to the nature of the environmental exposure in this population. Several ongoing collaborative studies involve tissue biomarkers of tumor behavior, aggressiveness, as well as whole genome analyses on cancer and normal adjacent tissues to understand if WTC exposure played a role in cancer etiology and outcome. We propose to update the existing biobank of human solid organ tissues from each cancer diagnosed after 2018, the date of last follow-up, among the WTC rescue and recovery workers, and to link the existing biobank to BioMe, an electronic medical record-linked blood biobank at MSSM of all patients' encounters who signed a research consent. The scope is to create an overall picture of the WTC responder's health, including clinical data and blood samples pre-cancer diagnosis, to facilitate future clinical and translational studies on WTC cancer etiology, biology, and cancer outcome. This project proposes to update the central repository of cancer tissue samples and adjacent normal tissue from each solid cancer diagnosed among the WTC Health Program (WTCHP) participants. This repository has the capability to be linked with the main WTCHP data set containing clinical, epidemiological and exposure information, as well as with the peripheral blood sample collected from the participants at the time of inclusion in the WTCHP. Through BioMe we will also access complete medical record history, and blood samples collected before or after a cancer diagnosis for WTC responders with cancer who are also MSSM patients. Clinical information collected by BioMe, and ancestry, will be available as well. Adding blood to the WTC cancer sample biobank will open the possibility of studies on temporal changes in systemic inflammation, metal and other toxicant exposure, metabolomics, and other biological markers over time. Blood-based markers can also be compared against markers observed in cancer tissue samples. We establish the tissue bank as a resource for the science community, by defining a process for qualified applicants to request available samples for research use, and for linking with the epidemiologic and clinical information in the database. A plan for the evaluation of the tissue bank usage and success, as measured by publications and funding obtained by investigators using the samples, is also included.

NIH Research Projects · FY 2026 · 2025-07

Project Summary Intact salivary glands facilitate several fundamental human needs, including breathing, swallowing, and lingual communication. However, salivary glands are extremely sensitive to irradiation (IR), and their critical functions are often irreversibly compromised following cancer radiation therapies. Previous attempts have identified mechanisms that could prevent IR damage and/or regenerate salivary epithelium in murine models. However, many of these pathways also promote cancer growth post-IR, significantly limiting their selectivity. Therefore, identifying mechanisms that can specifically protect or regenerate salivary glands without benefiting cancer growth post-IR remains a crucial scientific challenge that lacks efficient models. To solve this problem, we developed a scalable and rapid gene targeting system that can test over 2,600 genetic conditions in the salivary glands of a single mouse, significantly improving the efficiency of the current “one mouse one condition” paradigm of genetic modeling. Using this robust platform and a TetOn-shRNA pool, we tested 255 potential regulators of salivary epithelium growth for their ability to promote clonal expansion in salivary glands and syngeneic OSCC grafts post-IR. Contrasting these two sets of screens led to the identification of Smarca2 depletion as the top promoter of salivary gland-specific expansion post-IR without supporting cancer growth, standing out from the majority of hits shared by both tissues. In long-term observation, we found that Smarca2 depletion specifically promoted salivary epithelium growth by expanding the acinar population, which is particularly hard to replenish post-IR and represents a key area the field is trying to address. Pre-IR induction of shSmarca2 suggested that Smarca2 depletion did not prevent salivary epithelium from DNA damage, apoptosis, or senescence. While post-IR induction of shSmarca2 showed that Smarca2 depletion robustly increased the dividing population and their division potential in acinar cells during early-stage regeneration. Mechanistically, depleting Smarca2, a core component of the SWI/SNF chromatin remodeler complex, resulted in the shutdown of key terminal differentiation genes enriched for Smarca2 occupancy. Additionally, Smarca2 depletion drove robust indirect activation of regeneration genes, potentially through retargeting SWI/SNF or other remodelers. Intriguingly, this program was uncoupled from Smarca2 depletion in OSCC cells, potentially blocked by the cancer-specific epigenetic landscape. In the current project we will explore the cellular strategies of Smarca2 depletion driven salivary epithelium expansion and underlying chromatin remodeling mechanisms using both murine/human salivary gland and cancer models. Our discovery of Smarca2 depletion-driven acinar cell expansion represents a salivary gland-specific regeneration switch without promoting cancer growth. Accomplishing this project will establish a paradigm for understanding the distinct post-IR growth control mechanisms between salivary epithelium and cancer at the chromatin level, an area that is significantly underexplored.

NIH Research Projects · FY 2025 · 2025-07

Project Summary: The cohort of rescue and recovery workers (responders) involved in the emergency response and cleanup following the 2001 World Trade Center (WTC) disaster is aging (current median age is 60 years). Chronological aging is itself a major driver of disease. Aging, though, is a complex and nuanced biological process, and WTC responders may be experiencing “premature” or “accelerated” aging, whereby the body is aging faster than expected. Given the excess age-related morbidity and mortality associated with WTC exposure, including increased cancer risk, studies investigating WTC-associated premature aging on the biological level are warranted. Epigenetic clocks use DNA methylation levels at CpG sites throughout the genome to estimate biological age. An estimated biological age greater than actual chronological age reflects accelerated aging. In prior projects, our group has shown that DNA methylation was dysregulated because of WTC exposure, and that WTC exposure accelerated biological aging among women who are WTC survivors. Because sex is an effect measure modifier of both DNA methylation profiles and the normal aging process, studies are needed that explore this relationship in WTC-exposed males, including WTC rescue and recovery workers, who likely had higher levels of acute WTC exposure. Moreover, while DNA methylation markers derived from blood have been shown to mirror methylation signatures in tumor tissues, no studies have yet to validate tumor-tissue deregulated aging in responders. We propose here a study of WTC general responders (“exposed”) and males from the general population (“unexposed”). For each group we will calculate the mean biological age using validated epigenetic clock estimation techniques, including Horvath, Hannum, PhenoAge and GrimAge, and DunedinPACE. We will then compare rates of accelerated aging of WTC exposed vs unexposed participants, adjusting for important confounders. We intend to do this using both blood (Aim 1) and prostate tumor tissue (Aim 2). We hypothesize WTC exposure promotes accelerated aging among responders. We additionally expect to see similar aging-associated DNA methylation changes in both the blood of WTC cancer-free responders, as well as WTC tumor tissues. Improved understanding of how WTC exposure modifies the biological aging processes could improve both preventive and therapeutic care for responders, informing aging appropriate interventions such as exercise, nutritional supplementation, vitamin D, cognitive training, behavioral therapy, as a few examples. Biomarkers of dysregulated aging can also be used to screen for age-related vulnerability and conditions.

NIH Research Projects · FY 2025 · 2025-07

PROJECT SUMMARY Members of the SLC4 bicarbonate transporter family regulate cellular pH in essentially all tissues. Among those, SLC4A7, SLC4A8, and SLC4A10 constitute the subfamily of electroneutral, Na+-dependent transporters that are highly expressed in the brain. Control of cellular pH is not only important for essentially all cellular processes, but it uniquely also directly governs neuronal excitability, making SLC4A7, SLC4A8, and SLC4A10 key regulators of neuronal signaling. Unsurprisingly, mutations of these transporters thus cause a variety of neurological and neurodevelopmental disorders, and, for instance, loss of SLC4A10 function has been found to cause intellectual impairment, anxiety, hyperactivity, and episodic seizures in children. Although these clinical findings and studies in animal models highlight the importance of these transporters in human health and disease, little is known about their molecular mechanisms. Moreover, no tool compounds are currently available to further interrogate the function of SLC4A7, SLC4A8, and SLC4A10, and potentially explore therapeutic avenues in seizure disorders and other illnesses. To address these shortcomings we herein propose a multidisciplinary approach to combine insights from structural and biochemical studies, molecular dynamics simulations, structure-based in silico drug development, and validation of probe activity in (patho)physiologically relevant model systems. Our ultimate goal is to provide key mechanistic insights into SLC4 substrate binding, transport and inhibition, and directly leverage these data in the development of useful SLC4 tool compounds with different subtype-selectivity profiles. We specifically propose three independent aims: (i) Elucidate the different substrate stoichiometries of SLC4A7, SLC4A8, and SLC4A10 using a combination of cryoEM, computational approaches, and cellular transport studies; (ii) develop subtype- selective and nonselective SLC4 tool compounds targeting substrate and allosteric binding sites via structure- based computational drug discovery; and (iii) validate utility of novel probe scaffolds in primary neuronal culture, human neurons derived from induced pluripotent stem cells, and brain slices using substrate uptake assays and electrophysiological recordings. Together, our studies are poised to not only provide direct fundamental insights into SLC4 structure and function, but also generate first-in-class probes with which to explore therapeutic avenues at SLC4 in future studies.

NIH Research Projects · FY 2025 · 2025-07

Project Summary/Abstract People with Down Syndrome have many co-occurring health conditions, but one of the most common is Alzheimer’s Disease. People with Down Syndrome begin to accumulate Alzheimer’s pathology at much earlier ages than people without Down Syndrome. Accordingly, by the age of 60, almost all people with DS will be symptomatic for Alzheimer’s Disease dementia, whereas less than 10% of people without Down Syndrome will have Alzheimer’s Disease dementia. Because Alzheimer’s Disease impacts virtually all people with Down Syndrome, developing interventions to block or slow the progression of Alzheimer’s Disease may be especially helpful for increasing the quality of life for people with Down Syndrome. Our previous work showed that elevated levels of follicle-stimulating hormone (FSH) enhance the genesis and progression of Alzheimer’s pathology in mouse models of Alzheimer’s Disease. This led us to wonder whether Down Syndrome might be associated with elevated levels of FSH. Here, we will first measure FSH throughout the lifespan in two mouse models of Down Syndrome and explore possible mechanisms by which FSH might be altered in Down Syndrome. In our published work, we also developed a first-in-class monoclonal antibody targeting FSH; the antibody binds to the FSH receptor thereby preventing FSH from acting on its’ receptor. We will then determine whether administration of this antibody late in the lifespan reduces age-related accumulation of Alzheimer’s-like pathology in mouse models of Down Syndrome, as well as cognitive function. Lastly, we will compare antibody administration in young and aged mice with Down Syndrome and examine the impact on metabolic and non-metabolic conditions that are highly prevalent in Down Syndrome and arise from FSH- sensitive organs. The overarching goal of our proposed work is to determine if and when FSH is elevated in Down Syndrome, and probe whether reducing the actions of FSH helps normalize physiological function, thereby unmasking a new therapeutic target for the treatment of multiple co-occurring diseases in Down Syndrome.

NIH Research Projects · FY 2025 · 2025-07

PROJECT SUMMARY Salivary glands (SGs) are complex structures composed of heterogenous epithelial populations, each possessing unique and crucial functions that contribute to the production and secretion of saliva. Hyposalivation, caused by irreversible damage to SG epithelium in conditions such as Sjögren’s Syndrome or irradiation treatment for head and neck cancer, leads to chronic dry mouth and increased risk of dental caries, oral infections, and overall discomfort. The differentiation and organization of epithelial lineages during embryonic SG development relies on the surrounding niche, actively signaling to epithelial cells from the start of SG initiation and continuing into maturity. Developmental roles of tissue-resident macrophages (TRMs) have been defined in the prostate and mammary glands, yet despite prominence in the SG, their functional contribution during SG morphogenesis remains unknown. My preliminary work utilizing transcriptional analysis, transgenic mouse models, and explant cultures has defined a significant upregulation of TRMs to the SG niche during embryonic development. I have further characterized an immune-epithelial crosstalk via ligand-receptor signaling, and aberrant SG morphogenesis induced by a timed TRM-depletion, pointing to the necessity of TRMs in the regulation of SG development. Thus, I hypothesize that processes of SG formation are regulated through diverse TRM functions. In AIM 1, I will determine the functional contribution of TRMs through conditional depletions at multiple timepoints of early development and characterize changes to organization of major niche structures, SG epithelial morphology and differentiation dynamics, and global transcriptomic changes. In AIM 2, I will define the role of a secreted ligand signaling pathway in mediating SG epithelial and niche cell identity and differentiation dynamics through interrogation of single cell transcriptomics generated from transgenic global knockout mouse SGs, and leverage in vitro explant cultures to determine downstream signaling mechanisms in epithelium. Outcomes of this study will define temporal dynamics of immune-epithelial interactions and their contribution to SG development, and further characterize candidate factors as regulators of morphogenic processes with therapeutic potential in regenerating damaged SGs. The proposed project, supported by the Icahn School of Medicine at Mount Sinai, will provide me rigorous training in in vivo transgenics, embryonic tissue preparation, high-resolution imaging, epithelial explant culture, and bulk/single-cell transcriptomics, thereby preparing me for a future career as an independent researcher studying developmental and stem cell biology.

NIH Research Projects · FY 2025 · 2025-07

Project Summary Considerable progress has been made in identifying highly penetrant genes for autism spectrum disorder (ASD), particularly those involving deleterious de novo mutations that are evolutionarily constrained and crucial for developmental processes. These mutations typically result in syndromic forms of ASD with severe symptoms, but they are rare and do not account for the majority of ASD cases. In contrast, identifying genes with a moderate effect size (MES), which may explain the more heritable, mild, and broader ASD spectrum, has been limited. This project aims to identify and characterize MES risk genes by integrating both rare and common genetic variation. Our central hypothesis is that MES genes significantly contribute to ASD liability and manifestation, particularly among individuals with a high genetic load of common variation. The central hypothesis will be tested by pursuing four specific aims: (1) Identify and characterize MES risk genes using rare variation stratified by common variation; (2) Identify MES risk genes on chromosome X using rare variation stratified by common variation; (3) Identify MES risk genes contributing to ASD co-occurring conditions; and (4) Cluster MES risk genes to identify biological pathways relevant to ASD heterogeneity. This research is significant because it sheds light on the role of MES risk genes in ASD liability and presentation, enhancing diagnostic accuracy and supporting clinical applications such as genetic counseling and individualized treatment options. The most innovative parts of our proposal include (1) a novel gene discovery approach targeting MES risk genes typically overlooked due to their subtler effects, (2) the integration of both common and rare genetic variations to comprehensively understand their interplay in ASD, (3) investigating sex- and genetic ancestry-specific ASD risk factors, particularly on chromosome X, and (4) identifying genetic factors that correspond to the heterogeneity within ASD. Importantly, although methodologically innovative, we use large, established collections of individuals diagnosed with ASD, their family members, and population controls, re-purposing these datasets to produce novel and clinically useful findings.

NIH Research Projects · FY 2026 · 2025-07

ABSTRACT. Deficiencies in social recognition are common in psychiatric disorders such as schizophrenia and autism spectrum disorder. Effective treatments for these deficits are scarce, largely due to limited understanding of the brain circuitry governing social recognition memory. Gaining a comprehensive understanding of this circuitry is crucial for advancing our knowledge of this specific type of memory and pinpointing potential targets for treatment. The supramammillary nucleus (SuM) of the hypothalamus is a critical brain region that integrates spatial and social information. Our preliminary findings show decreased SuM activity during social interaction and the inhibition of SuM neurons projecting to the hippocampal CA2 region has been shown to play a pivotal role in promoting social novelty. Together, these results suggest a significant role of SuMàCA2 neurons in social recognition memory. However, the cellular, synaptic, and circuit mechanisms within the SuM that drive the SuMàCA2 neurons’ inhibition and memory formation, remain elusive. Social recognition memory is modulated by the oxytocin (OT) neuropeptide. Previous research has shown an abundance of OT fibers and OT receptors (OTRs) in the SuM, and recently we demonstrated that OTRs activity within the SuM is necessary for social recognition memory in rats. However, it remains unknown whether and how OT affects activity of SuM OTR- expressing neurons (SuM-OTR+) and SuMàCA2 projection neurons to facilitate social recognition memory. This proposal aims to address these key knowledge gaps by investigating the cellular and synaptic (Aim 1), as well as circuit mechanisms (Aim 2) through which OTR activation and SuM-OTR+ neurons impact the activity of SuMàCA2 projecting neurons to drive the formation of social recognition memory. Based on the scientific premise and our preliminary data, our overarching hypothesis proposes that by exciting SuM-OTR+ interneurons, OTRs inhibit SuMàCA2 activity, thereby facilitating the acquisition of social recognition memory. To test this hypothesis, we will employ a novel OTR-Cre rat line combined with in vitro electrophysiology and peptide optogenetics to test if, by exciting SuM-OTR+ GABA-ergic interneurons, OTR indirectly inhibits SuMàCA2 projection neurons in vitro (Aim 1). We will also use projection-specific chemogenetics to directly examine whether activating SuM-OTR+ neurons facilitates the acquisition of social recognition memory and whether this effect is abolished by the concurrent activation of SuMàCA2 neurons. Additionally, we will apply fiber photometry in behaving rats, combined with manipulation of OTR activity, to test the hypothesis that OTR excites SuM-OTR+ neurons and inhibits SuMàCA2 neurons in vivo. Since SuM also projects to the dentate gyrus (DG), we will examine if and how OT influences SuMàDG neuron activity during social memory acquisition and retrieval (Aim 2). This proposal will elucidate the SuM’s role within the broader network of social recognition memory, uncovering the cellular and circuit mechanisms that drive this process. Therefore, it holds significant translational potential for addressing social deficits commonly observed in many neuropsychiatric disorders.

NIH Research Projects · FY 2026 · 2025-07

Project Summary Considerable progress has been made in identifying highly penetrant genes for autism spectrum disorder (ASD), particularly those involving deleterious de novo mutations that are evolutionarily constrained and crucial for developmental processes. These mutations typically result in syndromic forms of ASD with severe symptoms, but they are rare and do not account for the majority of ASD cases. In contrast, identifying genes with a moderate effect size (MES), which may explain the more heritable, mild, and broader ASD spectrum, has been limited. This project aims to identify and characterize MES risk genes by integrating both rare and common genetic variation. Our central hypothesis is that MES genes significantly contribute to ASD liability and manifestation, particularly among individuals with a high genetic load of common variation. The central hypothesis will be tested by pursuing four specific aims: (1) Identify and characterize MES risk genes using rare variation stratified by common variation; (2) Identify MES risk genes on chromosome X using rare variation stratified by common variation; (3) Identify MES risk genes contributing to ASD co-occurring conditions; and (4) Cluster MES risk genes to identify biological pathways relevant to ASD heterogeneity. This research is significant because it sheds light on the role of MES risk genes in ASD liability and presentation, enhancing diagnostic accuracy and supporting clinical applications such as genetic counseling and individualized treatment options. The most innovative parts of our proposal include (1) a novel gene discovery approach targeting MES risk genes typically overlooked due to their subtler effects, (2) the integration of both common and rare genetic variations to comprehensively understand their interplay in ASD, (3) investigating sex- and genetic ancestry-specific ASD risk factors, particularly on chromosome X, and (4) identifying genetic factors that correspond to the heterogeneity within ASD. Importantly, although methodologically innovative, we use large, established collections of individuals diagnosed with ASD, their family members, and population controls, re-purposing these datasets to produce novel and clinically useful findings.

NIH Research Projects · FY 2025 · 2025-07

The liver, metabolic syndrome, and increased CVD risk among non-Hispanic Black World Trade Center (WTC) General Responders. This discovery-oriented R21 project has the following premise: In addition to tra- ditional cardiometabolic risk factors, upstream drivers of cardiovascular disease (CVD) in members of the WTC General Responder Cohort include: Aim I: Pleiotropic genetic variants that effect both liver metabolism and CVD, and Aim II: Social determinants of health (SDoH). The proposed studies address a critical problem. More intense exposure to the WTC dust cloud increases CVD. CVD diagnoses are especially high in non-Hispanic (NH) Black female WTC General Responders and mortality from CVD is about 2-fold higher in NH Black than in NH White WTC rescue and recovery workers. CVD often develops in the setting of a systemic abnormality called cardio- vascular-kidney metabolic syndrome (CKM-S). Liver steatosis and MASLD are the hepatic manifestations of CKM-S. CKM-S can lead to oxidative stress and altered liver metabolism. These changes affect CVD risk, as the liver produces key CVD risk modifiers (LDL, HDL triglycerides). Our goal is to achieve a deeper understand- ing of CVD drivers, including the genomic architecture of liver-related CVD risk factors and actionable SDoH. Aim I. Hypothesis: Pleiotropic genetic variants that affect MASLD risk (up or down) and that increase CVD risk have a higher frequency in NH Black General Responders and other NH Black populations than in NH Whites. Approach: To test the hypothesis and investigate the pathways and clinical consequences associated with these variants we will: [1] Analyze results of publicly available GWAS studies that identify single nucleotide polymor- phisms (SNPs) associated with MASLD and CVD and select SNPs with genome-wide significant pleiotropic signals, i.e., significant effect sizes (βs) for both liver and cardiovascular abnormalities; [2] Determine the burden of these variants across populations and use them to estimate a Polygenic Score (PGS) to test the association between their cumulative effect and clinical phenotypes in two large multiethnic biobanks, BioMe and All of Us cohorts. [3] Analyze the risk alleles and apply the PGS to data of WTC General Responders enrolled in BioMe. Aim II. Hypothesis: NH Black General Responders have a higher burden of SDoH than NH White General Re- sponders and SDoH contribute to disparities in CVD risk, as estimated by PREVENT equations of the American Heart Association (AHA); to increased CVD events; and to early age of CVD on-set. Approach: [1] Differences in the burden of SDoH will be compared between NH Black and NH White General Responders, both men and women; [2] CVD risk will be estimated using AHA PREVENT equations and compared among groups, with adjustment for age at the time of risk calculation; [3] If differences in CVD risk are found, the percentage of the disparity mediated by SDoH will be determined; [4] Hazard ratios for CVD diagnosis (total and by CVD subtype) and the age at first CVD diagnosis will be compared between NH Black and NH White General Responders and the percentage of any disparity mediated by SDoH will be determined for men and women.

NIH Research Projects · FY 2025 · 2025-06

PROJECT SUMMARY/ABSTRACT The past decade has witnessed groundbreaking work associating clonal hematopoiesis (CH), where acquired mutations define clonal outgrowths of hematopoietic stem cells, with chronic inflammatory diseases including solid cancers. While we have begun to understand suppressive mechanisms that hinder antitumor immunity, it remains unclear how immune cells in the tumor microenvironment behave differently when originating from progenitors carrying CH mutations like DNMT3A. Recent studies have established myeloid-biased hematopoiesis in the context of both DNMT3A CH and cancer-induced emergency myelopoiesis. Mechanistically, our laboratory has suggested the ability of lung tumors to communicate with distal bone marrow niches, pushing immune progenitors to produce corrupted progeny. Furthermore, recent work has shown that tumors can epigenetically rewire myeloid progenitors to increase production of immunosuppressive macrophages and interfere with tumor control. Expanding on these studies, I hypothesize that hematopoietic progenitors are molecularly reprogrammed by CH-related DNMT3A, increasing myeloid-driven immunosuppression, and enhancing bone marrow dysfunction in response to tumor-associated inflammation. Aim 1 will characterize the epigenetic wound inflicted by lung adenocarcinoma through a cohort of progenitor-enriched peripheral blood samples from lung cancer patients with DNMT3A CH. Using integrated single-cell multiomics and genotyping, I will trace molecular changes driven by DNMT3A mutation along the lineage of immune hematopoietic progenitors to circulating monocytes. I will assess the contribution of uncovered pathways to driving inflammatory phenotypes in DNMT3A mutated cell line experiments. Aim 2 extends the investigation to the lung tumors of DNMT3A CH patients, through spatial transcriptomics and genotyping. Spatial methods do not require tissue dissociation, which enables localization of both tumor myeloid cells arising from DNMT3A-mutated hematopoietic progenitors and wildtype cells. Genome-wide transcriptomic readouts will capture the full effects of enhanced immunoregulatory signaling and CH-associated inflammation on tumor immune responses. I will use co-cultures of lung cancer spheroids and myeloid cells as a simplified tumor microenvironment model to interrogate spatial transcriptomic findings. Together, these aims will identify CH-associated molecular programs in hematopoietic progenitors which may serve as novel neoadjuvant targets in patients refractory to conventional therapy. Leveraging the excellent training environment at Mount Sinai, this proposal bridges computational analyses, biological discovery, and clinical potential, serving as a well-rounded foundation for my career as an academic hematologist/oncologist.

NIH Research Projects · FY 2025 · 2025-06

Project Summary A plethora of studies over the past 30+ years have demonstrated that lack of proficient epigenetic regulation contributes significantly to neurodevelopmental disorders (NDDs). Particularly, histone H3 lysine 4 tri-methylation (H3K4me3) regulation has been found to be aberrantly regulated in multiple NDDs, including autism spectrum disorders (ASD). Of note, dysregulation of H3K4me3 domains was found in post-mortem brain of ASD patients, with spreading of “broad” domains (>5 kb) noted. H3K4me3 broad domains have previously been linked to expression of cell type specific genes, such as those involved in synaptic signaling. This spreading has been linked to lysine methyltransferase 2e (KMT2E, or MLL5), which contains a plant homeobox domain (PHD) that binds to H3K4me3, as well as a SET domain (lacking methyltransferase activity). Over 60 individual patients have been identified with heterozygous mutations in Kmt2e, who have symptoms related to intellectual disability, developmental delay, and epilepsy. H3K4me3 is adjacent to H3 glutamine 5, which can be serotonylated (H3Q5ser), creating a dual modification H3K4me3Q5ser. This combinatorial mark has been shown to enhance permissive transcription of target genes compared to H3K4me3 alone. Preliminary data shows H3K4me3Q5ser enhances significantly the binding of MLL5PHD versus H3K4me3 alone. ChIP-sequencing of H3K4me3Q5ser in embryonic forebrain tissues identified ~4000 H3K4me3Q5ser broad domains which are enriched at genes involved in neurodevelopment-associated processes. Kmt2e-/- mice exhibit significant behavioral alterations, which match known developmental defect phenotypes of patients with KMT2E mutations. Immunoprecipitation- mass spectrometry of MLL5 revealed strong interactions with the nuclear co-receptor/histone deacetylase 3 (NCOR/HDAC3) complex, as well as lysine demethylase 5a (KDM5A, or JARID1A), which regulate H3/H4ac and H3K4me3 respectively. Thus, I hypothesize that H3K4me3Q5ser recruits MLL5 to key developmental loci during neurodevelopment, secondarily recruiting the NCOR/HDAC3 complex and JARID1A, regulating H3K4me3Q5ser broad domains, allowing for appropriate transcriptional programming in brain. First, I will characterize the molecular interactions between H3K4me3Q5ser, MLL5, NCOR/HDAC3, and JARID1A in cellulo, assessing key protein domains for direct protein:protein interactions and the requirement of MLL5 for co-targeting genome wide. Secondly, using our Kmt2e-/- mouse, I will assess the impact of MLL5 loss on histone modification landscapes in the developing brain, across both embryonic and postnatal development and intersecting this with transcriptomic profiling. Finally, I will assess the impact of Kmt2e on neurodevelopment by attempting to virally ‘rescue’ proper gene expression in Kmt2e-/- mice during postnatal development using viral expression, measuring changes in H3K4me3Q5ser levels and behavioral phenotypes versus control animals. Together, these aims will provide valuable insights into this novel epigenetic mechanism which regulates neurodevelopment, which is perturbed in patients with NDDs which have mutations in one of the aforementioned genes.

NIH Research Projects · FY 2025 · 2025-06

PROJECT SUMMARY Multiple Myeloma (MM) is an incurable plasma cell malignancy and is the second most common hematologic malignancy. SETD8 (also known as SET8, PR-Set7, or KMT5A) is the lysine methyltransferase that is responsible for monomethylation of histone H4 lysine 20 (H4K20me), an important histone methylation mark. SETD8 also monomethylates non-histone substrates such as the tumor suppressor p53 and replication factor PCNA, causing suppression of p53-dependent transcriptional activation in cancer cells and promoting cancer cell proliferation. Consistent with this, SETD8 is overexpressed in numerous cancers. In particular, we recently reported that SETD8 is overexpressed in relapsed primary MM and high SETD8 expression is associated with poor prognosis. Importantly, SETD8 knockdown (KD) effectively suppressed the proliferation of SETD8-high MM cells, but not SETD8-low MM cells and normal cells. Moreover, primary malignant plasma cells are particularly addicted to the methyltransferase activity of SETD8. We therefore hypothesized that pharmacological inhibition of SETD8 could be a novel and effective therapeutic strategy for the treatment of MM. In our preliminary studies, we discovered UNC0379 (the first SETD8 selective inhibitor), MS453 (the first SETD8 selective covalent inhibitor), and MS2928 (the most potent and selective SETD8 inhibitor to date). Inhibition of SETD8 by UNC0379 reduced H4K20me, induced cell-cycle defects and apoptosis, and suppressed the growth of MM cell lines and primary MM cells without significant toxicity in non-myeloma cells. Furthermore, MS2928 reduced H4K20me more effectively than UNC0379 and effectively inhibited the growth of SETD8-high MM cells, but not SETD8-low MM cells and normal cells, thereby phenocopying the effect of SETD8 KD. Importantly, MS2928 was bioavailable in mouse pharmacokinetic studies and significantly inhibited tumor growth in vivo in two MM cell line xenograft mouse models without apparent toxicity. Encouraged by these promising preliminary results, we propose to optimize our SETD8 inhibitors into in vivo chemical probes and evaluate SETD8 selective inhibitors in MM cellular and mouse models to further test and validate our therapeutic hypothesis. The optimized SETD8 inhibitors to be generated in this project will also be invaluable chemical tools for assessing the therapeutic potential of SETD8 inhibition in other SETD8-overexpressing cancers, and can be further optimized into drug candidates in the future and ultimately translated in the clinic for cancer patients.